Preclinical Trials for Prevention of Tumor Progression of Hepatocellular Carcinoma by LZ-8 Targeting c-Met Dependent and Independent Pathways

Abstract

Hepatocellular carcinoma (HCC) is among the most lethal cancers. Mounting studies highlighted the essential role of the HGF/c-MET axis in driving HCC tumor progression. Therefore, c-Met is a potential therapeutic target for HCC. However, several concerns remain unresolved in c-Met targeting. First, the status of active c-Met in HCC must be screened to determine patients suitable for therapy. Second, resistance and side effects have been observed frequently when using conventional c-Met inhibitors. Thus, a preclinical system for screening the status of c-Met signaling and identifying efficient and safe anti-HCC agents is urgently required. In this study, immunohistochemical staining of phosphorylated c-Met (Tyr1234) on tissue sections indicated that HCCs with positive c-Met signaling accounted for approximately 46% in 26 cases. Second, many patient-derived HCC cell lines were established and characterized according to motility and c-Met signaling status. Moreover, LZ8, a medicinal peptide purified from the herb Lingzhi, featuring immunomodulatory and anticancer properties, was capable of suppressing cell migration and slightly reducing the survival rate of both c-Met positive and negative HCCs, HCC372, and HCC329, respectively. LZ8 also suppressed the intrahepatic metastasis of HCC329 in SCID mice. On the molecular level, LZ8 suppressed the expression of c-Met and phosphorylation of c-Met, ERK and AKT in HCC372, and suppressed the phosphorylation of JNK, ERK, and AKT in HCC329. According to receptor array screening, the major receptor tyrosine kinase activated in HCC329 was found to be the epidermal growth factor receptor (EGFR). Moreover, tyrosine-phosphorylated EGFR (the active EGFR) was greatly suppressed in HCC329 by LZ8 treatment. In addition, LZ8 blocked HGF-induced cell migration and c-Met-dependent signaling in HepG2. In summary, we designed a preclinical trial using LZ8 to prevent the tumor progression of patient-derived HCCs with c-Met-positive or -negative signaling.

Induction of Cbl-dependent epidermal growth factor receptor degradation in Ling Zhi-8 suppressed lung cancer

“…Here, we showed that the rLZ‐8 induced the Cbl‐mediated clathrin‐dependent degradation of both wt‐ and mut‐EGFR and also exhibited efficacy against lung cancer cells. In contrast, rLZ‐8 did not downregulate other membrane proteins, including transforming growth factor β (TGFβ) receptors, fibroblast growth factor receptor (FGFR) and integrin beta 1, but also induced c‐Met (hepatocyte growth factor receptor) degradation as previous study (data not shown). Moreover, our unpublished data showed that rLZ‐8 also inhibits cell viability and suppresses EGFR expression in various cancer cells, such as breast, head and neck, colon and cervical cancer (Supplementary Fig.…”

Hydrogen peroxide inducible clone-5 mediates reactive oxygen species signaling for hepatocellular carcinoma progression

“…Initially, we compared the expression of Hic-5 in several patient-derived HCC cell lines, the phenotypes of which such as cell motility have been characterized [ 20 ]. As shown in Figure 2A , Hic-5 markedly expressed in most of the HCCs such as HCC329, HCC353, HCC365 and HCC372 with average to high motility, slightly expressed in HCC363 with average motility but not in the non-motile HCC340 and HCC374 [ 20 ]. Notably, HCC329, the most motile HCC [ 20 ], expressed the highest Hic-5 among the HCCs examined.…”

“…As shown in Figure 2A , Hic-5 markedly expressed in most of the HCCs such as HCC329, HCC353, HCC365 and HCC372 with average to high motility, slightly expressed in HCC363 with average motility but not in the non-motile HCC340 and HCC374 [ 20 ]. Notably, HCC329, the most motile HCC [ 20 ], expressed the highest Hic-5 among the HCCs examined.…”

“…Thus, Hic-5 was essential for motility and invasiveness of the HCC. Further, the role of Hic-5 in metastatic capability of HCCs was investigated using the most motile HCC329 in a SCID mice model established previously [ 20 ]. The cells (as concentrated pellet) were injected into the middle lobe of liver followed by observation of the growth of primary tumor on middle lobe and metastatic loci on left or right lobe.…”

An Epitope on EGFR Loading Catastrophic Internalization Serve as a Novel Oncotarget for Hepatocellular Carcinoma Therapy

“…A protein (Lingzhi-8, LZ-8) from Ganoderma Lucidum was reported that it had anti-tumor activity and could modulate EGFR expression but its binding site on EGFR and mechanism are still unclear [18,19]. Here, we found that recombinant LZ-8 (rLZ-8) could bind to EGFR specifically, induce catastrophic macropinocytosis, enter into HCC cells with EGFR, lead to the blockage of cell membrane recycling and then result in cell death via membrane ruffling, cell membrane over-internalization, cell rounding and bursting.…”